Graft-vs-host disease (GVHD) is the major complication of hematopoietic stem cell therapies that are currently used to treat hematological malignancies, and is associated with very high rates of morbidity and mortality. Therefore, the development of new approaches to prevent or ameliorate this pathology would be of great clinical value. The studies proposed here will investigate whether human NKT cells, a population of innate T lymphocytes that has been shown to have potent immunoregulatory functions, can be used to inhibit the development of GVHD in a humanized mouse model system. In this model system, severely immune deficient mice are transplanted with human hematopoietic stem cells, along with fetal liver and thymic tissue. This results in the differentiation and subsequent long-term persistence of multiple lineages of human immune cells in the mice, including T cells, B cells, and myeloid cell types. At late time points after transplantation (i.e. greater than 100 days) the mice show signs of a condition that markedly resembles chronic GVHD in human hematopoietic transplant patients. We will use this in vivo model system to test the immunoregulatory impact of human NKT cells. By manipulating parameters such as whether CD4+ or CD4- NKT cells are the dominant subset, the strength of antigenic activation, the level of NKT cell activation by the endogenous ligand LPC, and the characteristics of CD1d+ APCs, we will gain insight into the factors that determine the immunoregulatory effects of NKT cells. The specific aims are as follows: Assess the impact of the following treatments on the development of GVHD: a) adoptively transferring in vitro expanded NKT cells (CD4+ vs. CD4- subsets); b) administering glycolipid antigens (strong vs. weak TCR agonists); c) augmenting or interrupting NKT cell recognition of LPC (an endogenous lipid antigen associated with inflammation); d) treatment with a PPAR? agonist that causes up-regulation of CD1d expression on myeloid APCs and alters their cytokine secretion to a less inflammatory profile. PUBLIC HEALTH RELEVANCE: Cancers of the immune system (leukemias, lymphomas, myelomas, etc.) can be successfully treated by transplanting bone marrow derived cells from a healthy person into the patient; however, a serious complication that frequently arises is that the new immune cells attack the patient. This grant will investigate using a small population of human lymphocytes called NKT cells to prevent this complication. Positive results from these studies could quickly open the way for implementing similar approaches in treating human cancers.